Abuse of methamphetamine (METH) and 3,4-methylenedioxymethamphetamine (MDMA) is increasing at a steady and alarming rate. Recent evidence has linked use of these drugs to persistent, if not permanent, neurochemical deficits in dopamine (DA) and serotonin (5HT) nerve endings. The DA transporter (DAT) and tyrosine hydroxylase (TH), and the 5HT transporter (5HTT) and trytophan hydroxylase (TPH) are phenotypic markers for DA and 5HT nerve endings, respectively. These proteins bear the brunt of damage associated with METH and MDMA. The coincident reduction in function of monoamine transporters and monooxygenase enzymes is significant because it leads to reductions in the synaptic and intracellular levels of DA and 5HT, and could have direct effects on neurotransmission in these neurochemical systems. In view of the potential for long-term neuronal deficits associated with these drugs, it is imperative that their toxic mechanisms of action be more fully understood. The long-term goal of this renewal application is to increase the understanding of how METH and MDMA cause damage to DA and 5HT nerve endings. The general, working hypothesis driving these two studies is that METH and MDMA transduce their toxic effects on nerve endings through reactive oxygen species and reactive nitrogen species. It is hypothesized that the interaction of nitric oxide and superoxide, two reactants already suspected as mediators of METH and MDMA toxicity, produces peroxynitrite (ONOO-). We hypothesize further that the obligatory role of DA in METH/MDMA toxicity will be revealed through a better understanding of its reaction with ONOO- to yield DA-quinone. Consequently, ONOO- and DA-quinone emerge as novel reactants whose participation in METH/MDMA actions has not been investigated heretofore. Specific Aim 1 will determine the molecular mechanisms by which ONOO- and DA-quinone diminish TH and TPH catalytic activity. Specific Aim 2 will investigate how these reactants affect DAT and 5HTT function (transport and ligand recognition site) in cells stably expressing these proteins. Specific Aim 3 will search for molecular footprints of ONOO- and DA-quinone action in monoamine nerve endings after treatment of mice with METH or MDMA. These studies will test novel hypotheses relating to the identification of reactive species that mediate METH and MDMA effects on monoamine nerve endings.